Simultaneous visualization of cysteine/homocysteine and glutathione in living cells and Daphnia magna via dual-signaling fluorescent chemosensor

Biological thiols, including cysteine (Cys), homocysteine (Hcy) and glutathione (GSH), play crucial roles in living organisms, such as maintaining the cellular redox equilibrium, and are closely related to many diseases. The development of efficient fluorescent probes that can diacritically detect biothiols with similar reactivity are of great desirable for the study of biological processes. The conventional single-emission fluorescent probes may be disturbed by surrounding environment and biological systems, which is greatly lose the accuracy of its detection. Herein, we have constructed a fluorescent probe (TCF-NBD) by connecting two fluorophores with a biothiol-reactive linker, which has two separated emissions at 547 nm and 610 nm via different excitations (470 nm and 570 nm, respectively), for simultaneously fast detecting Cys/Hcy and GSH with high selectivity and sensitivity. Furthermore, the probe TCF-NBD exhibits low cytotoxicity and was successfully applied to visualize intracellular biothiols in living cells and Daphnia magna by fluorescence imaging. We believe that TCF-NBD can be a powerful tool for conducting the research of thiol-related diseases.